The invention relates to compositions comprising an immunomodulatory molecule (IMM) including an antigen, conjugated to a polynucleotide that contains or consists of at least one immunostimulatory oligonucleotide (ISS-PN). It also relates to methods for modulating the immune response of a vertebrate host to an antigen.
Conventionally, immunization of a host against an antigen is accomplished by repeatedly vaccinating the host with the antigen. While most current vaccines elicit reasonable antibody responses, cellular responses (in particular, major histocompatibility complex (MHC) class I-restricted cytotoxic T cells) are generally absent or weak. For many infectious diseases, such as tuberculosis and malaria, humoral responses are of little protective value against infection.
Given the weak cellular immune response to protein antigens, modulation of the immune responses to these antigens has clear importance. The ability to modify immune responses to protein or peptide antigen has implications for tumor therapy, for the treatment of allergic disorders and for treatment of other conditions achievable through induction of a vigorous cellular immune response.
The present invention provides compositions comprising an ISS-PN which is conjugated to an IMM (which includes an antigen) to form ISS-PN/IMM conjugates. The ISS-PN/IMM conjugates of the invention are biological response modifiers in the sense that they modify the humoral and cellular immune response of a host to an antigen.
Specifically, the ISS-PN and IMM components of the ISS-PN/IMM conjugates synergistically boost the magnitude of the host immune response against an antigen to a level greater than the host immune response to either the IMM, antigen or ISS-PN alone. The ISS-PN/IMM conjugates also shift the host cellular immune response away from the helper T lymphocyte type 2 (Th2) phenotype toward a helper T lymphocyte type 1 (Th1) phenotype. These responses to ISS-PN/IMM conjugates are particularly acute during the important early phase of the host immune response to an antigen.
To these ends, ISS-PN/IMM conjugates are delivered by any route through which antigen-sensitized host tissues will be contacted with the ISS-PN/IMM conjugate. ISS-PN/IMM conjugates administered in this fashion boost both humoral (antibody) and cellular (Th1 type) immune responses of the host. Thus, use of the method to boost the immune responsiveness of a host to subsequent challenge by a sensitizing antigen without immunization avoids the risk of Th2-mediated, inmmunization-induced anaphylaxis by suppressing IgE production in response to the antigen challenge. An especially advantageous use for this aspect of the invention is treatment of localized allergic responses in target tissues where the allergens enter the body, such as the skin and mucosa.
Suppression of the Th2 phenotype according to the invention is also a useful in reducing antigen-stimulated IL-4 and IL-5 production. Thus, the invention encompasses delivery of ISS-PN/IMM conjugates to a host to suppress the Th2 phenotype associated with conventional antigen immunization (e.g., for vaccination or allergy immunotherapy).
The shift to a Th1 phenotype achieved according to the invention is accompanied by increased secretion of IFN xcex1, xcex2 and xcex3, as well as IL-12 and IL-18. Each of these cytokines enhance the host""s immune defenses against intracellular pathogens, such as viruses. Thus, the invention encompasses delivery of ISS-PN/IMM conjugates to a host to combat pathogenic infection.
Angiogenesis is also enhanced in the Th1 phenotype (ostensibly through stimulation by IL-12). Thus, the invention encompasses delivery of ISS-PN/IMM conjugates to a host to stimulate therapeutic angiogenesis to treat conditions in which localized blood flow plays a significant etiological role; e.g., retinopathies.
The ISS-PN/IMM conjugates of the invention comprise an IMM conjugated to a polynucleotide that includes, or consists of, at least one immunostimulatory oligonucleotide (ISS-ODN) moiety. The ISS-ODN moiety is a single- or double-stranded DNA or RNA oligonucleotide having at least 6 nucleotide bases which may include, or consist of, a modified oligonucleoside or a sequence of modified nucleosides.
The ISS-ODN moieties comprise, or may be flanked by, a CpG containing nucleotide sequence or a p(IC) nucleotide sequence, which may be palindromic. Where the oligonucleotide moiety comprises a CpG sequence, it may include a hexamer structure consisting of: 5xe2x80x2-Purine, Purine, CG, Pyrimidine, Pyrimidine-3xe2x80x2. Examples of such hexamer structures are AACGTT, AGCGTT, GACGTT, GGCGTT, AACGTC, and AGCGTC.
In one aspect of the invention, the ISS-PN consists of an ISS-ODN. Alternatively, the ISS-PN comprises an ISS-ODN.
Conjugates of the invention also include PN/IMM wherein the PN serves as a carrier to introduce the IMM antigen into MHC Class I processing pathways not normally stimulated by soluble antigen, but lacks ISS activity and therefore does not stimulate a Th1 phenotype immune response. Examples of such PN/IMM are those wherein the CpG motif is mutated, for example, to a GpG motif.
In one aspect of the invention, the IMM conjugate partner to the ISS-PN consists of an antigen. Such antigens are selected from the group of antigens consisting of proteins, peptides, glycoproteins, polysaccharides and gangliosides.
In another aspect of the invention, the IMM conjugate partner comprises an antigen and further comprises an immunostimulatory molecule selected from the group of such molecules consisting of adjuvants, hormones, growth factors, cytokines, chemokines, targeting protein ligands, and trans-activating factors.
In another aspect of the invention, the ISS-PN/IMM conjugate is modified for targeted delivery by, for example, attachment to a monoclonal antibody, receptor ligand and/or liposome.
Pharmaceutically acceptable compositions of ISS-PN/IMM conjugates are provided for use in practicing the methods of the invention. Where appropriate to the contemplated course of therapy, the ISS-PN/IMM conjugates may be administered with anti-inflammatory or immunotherapeutic agents. Thus, a particularly useful composition for use in practicing the method of the invention is one in which an anti-inflammatory agent (e.g., a glucocorticoid) is mixed with, or further conjugated to, an ISS-PN/IMM conjugate.
The ISS-PN/IMM conjugates can also be provided in the form of a kit comprising ISS-PN/IMM conjugates and any additional medicaments, as well as a device for delivery of the ISS-PN/IMM conjugates to a host tissue and reagents for determining the biological effect of the ISS-PN/IMM conjugates on a treated host.